Exposure to Low-Dose 56Fe-Ion Radiation Induces Long-Term Epigenetic Alterations in Mouse Bone Marrow Hematopoietic Progenitor and Stem Cells

Abstract Low-toxicity conditioning regimens prior to bone marrow transplantation (BMT) are widely explored. We developed a new protocol using hematopoietic growth factors prior to low-dose total body irradiation (TBI) in recipients of autologous transplants to establish high levels of long-term donor cell engraftment. We hypothesized that treatment of recipient mice with growth factors would selectively deplete stem cells, resulting in successful long-term donor cell engraftment after transplantation. Recipient mice were treated for 1 or 7 days with growth factors (stem cell factor [SCF] plus interleukin 11 [IL-11], SCF plus Flt-3 ligand [FL], or granulocyte colony-stimulating factor [G-CSF]) prior to low-dose TBI (4 Gy). Donor cell chimerism was measured after transplantation of congenic bone marrow cells. High levels of donor cell engraftment were observed in recipients pretreated for 7 days with SCF plus IL-11 or SCF plus FL. Although 1-day pretreatments with these cytokines initially resulted in reduced donor cell engraftment, a continuous increase in time was observed, finally resulting in highly significantly increased levels of donor cell contribution. In contrast, G-CSF treatment showed no beneficial effects on long-term engraftment. In vitro stem cell assays demonstrated the effect of cytokine treatment on stem cell numbers. Donor cell engraftment and number of remaining recipient stem cells after TBI were strongly inversely correlated, except for groups treated for 1 day with SCF plus IL-11 or SCF plus FL. We conclude that long-term donor cell engraftment can be strongly augmented by treatment of recipient mice prior to low-dose TBI with hematopoietic growth factors that act on primitive cells.

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Efficient retroviral gene transfer to purified long-term repopulating hematopoietic stem cells

Blood ◽

10.1182/blood.v84.1.74.74 ◽

1994 ◽

Vol 84 (1) ◽

pp. 74-83 ◽

Cited By ~ 36

Author(s):

SJ Szilvassy ◽

S Cory

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Cell Biology ◽

Bone Marrow Cells ◽

High Titer ◽

Marker Gene ◽

Mouse Bone Marrow ◽

Hematopoietic Stem

Abstract Efficient gene delivery to multipotential hematopoietic stem cells would greatly facilitate the development of effective gene therapy for certain hematopoietic disorders. We have recently described a rapid multiparameter sorting procedure for significantly enriching stem cells with competitive long-term lymphomyeloid repopulating ability (CRU) from 5-fluorouracil (5-FU)-treated mouse bone marrow. The sorted cells have now been tested as targets for retrovirus-mediated delivery of a marker gene, NeoR. They were cocultured for 4 days with fibroblasts producing a high titer of retrovirus in medium containing combinations of the hematopoietic growth factors interleukin-3 (IL-3), IL-6, c-kit ligand (KL), and leukemia inhibitory factor (LIF) and then injected into lethally irradiated recipients, together with sufficient “compromised” bone marrow cells to provide short-term support. Over 80% of the transplanted mice displayed high levels (> or = 20%) of donor- derived leukocytes when analyzed 4 to 6 months later. Proviral DNA was detected in 87% of these animals and, in half of them, the majority of the hematopoietic cells were marked. Thus, infection of the stem cells was most effective. The tissue and cellular distribution of greater than 100 unique clones in 55 mice showed that most sorted stem cells had lymphoid as well as myeloid repopulating potential. Secondary transplantation provided strong evidence for infection of very primitive stem cells because, in several instances, different secondary recipients displayed in their marrow, spleen, thymus and day 14 spleen colony-forming cells the same proviral integration pattern as the primary recipient. Neither primary engraftment nor marking efficiency varied for stem cells cultured in IL-3 + IL-6, IL-3 + IL-6 + KL, IL-3 + IL-6 + LIF, or all four factors, but those cultured in IL-3 + IL-6 + LIF appeared to have lower secondary engraftment potential. Provirus expression was detected in 72% of the strongly marked mice, albeit often at low levels. Highly efficient retroviral marking of purified lymphomyeloid repopulating stem cells should enhance studies of stem cell biology and facilitate analysis of genes controlling hematopoietic differentiation and transformation.

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Isolation and Assessment of Long‐Term Reconstituting Hematopoietic Stem Cells from Adult Mouse Bone Marrow

Current Protocols in Stem Cell Biology ◽

10.1002/9780470151808.sc02a04s3 ◽

2007 ◽

Vol 3 (1) ◽

Cited By ~ 4

Author(s):

David Kent ◽

Brad Dykstra ◽

Connie Eaves

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Adult Mouse ◽

Mouse Bone Marrow ◽

Hematopoietic Stem

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Differentiation pathways of pluripotent hemopoietic stem cells in long-term mouse bone marrow culture

Leukemia Research ◽

10.1016/0145-2126(86)90032-9 ◽

1986 ◽

Vol 10 (3) ◽

pp. 345-351

Author(s):

Hiroko Izumi ◽

Martine Guigon ◽

Emilia Frindel

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Bone Marrow Culture ◽

Mouse Bone Marrow ◽

Hemopoietic Stem Cells ◽

Differentiation Pathways

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Diabetes impairs the interactions between long-term hematopoietic stem cells and osteopontin-positive cells in the endosteal niche of mouse bone marrow

AJP Cell Physiology ◽

10.1152/ajpcell.00400.2012 ◽

2013 ◽

Vol 305 (7) ◽

pp. C693-C703 ◽

Cited By ~ 14

Author(s):

Hironori Chiba ◽

Koji Ataka ◽

Kousuke Iba ◽

Kanna Nagaishi ◽

Toshihiko Yamashita ◽

...

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Mouse Bone Marrow ◽

Diabetic Mice ◽

Hematopoietic Stem ◽

Coculture System ◽

Endosteal Niche

Hematopoietic stem cells (HSCs) are maintained, and their division/proliferation and quiescence are regulated in the microenvironments, niches, in the bone marrow. Although diabetes is known to induce abnormalities in HSC mobilization and proliferation through chemokine and chemokine receptors, little is known about the interaction between long-term HSCs (LT-HSCs) and osteopontin-positive (OPN) cells in endosteal niche. To examine this interaction, LT-HSCs and OPN cells were isolated from streptozotocin-induced diabetic and nondiabetic mice. In diabetic mice, we observed a reduction in the number of LT-HSCs and OPN cells and impaired expression of Tie2, β-catenin, and N-cadherin on LT-HSCs and β1-integrin, β-catenin, angiopoietin-1, and CXCL12 on OPN cells. In an in vitro coculture system, LT-HSCs isolated from nondiabetic mice exposed to diabetic OPN cells showed abnormal mRNA expression levels of Tie2 and N-cadherin. Conversely, in LT-HSCs derived from diabetic mice exposed to nondiabetic OPN cells, the decreased mRNA expressions of Tie2, β-catenin, and N-cadherin were restored to normal levels. The effects of diabetic or nondiabetic OPN cells on LT-HSCs shown in this coculture system were confirmed by the coinjection of LT-HSCs and OPN cells into bone marrow of irradiated nondiabetic mice. Our results provide new insight into the treatment of diabetes-induced LT-HSC abnormalities and suggest that the replacement of OPN cells may represent a novel treatment strategy.

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Hematopoiesis and stem cell renewal in long-term bone marrow cultures containing catalase

Blood ◽

10.1182/blood-2005-03-1180 ◽

2006 ◽

Vol 107 (5) ◽

pp. 1837-1846 ◽

Cited By ~ 28

Author(s):

Rashmi Gupta ◽

Simon Karpatkin ◽

Ross S. Basch

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Ex Vivo ◽

Mouse Bone Marrow ◽

Cell Renewal ◽

Hematopoietic Stem ◽

Stem Cell Renewal

Culturing mouse bone marrow in the presence of catalase dramatically alters hematopoiesis. Granulocyte output is initially increased 4- to 5-fold. This increase is transient and granulocyte production declines as immature (Sca-1+/LIN-) cells accumulate. One third of these immature cells have a phenotype (Sca-1+/c-Kit+) characteristic of hematopoietic stem cells. At 2 to 3 weeks there are greater than 200-fold more Sca-1+/c-Kit+/LIN- cells in treated cultures than in controls. This population contains functional stem cells with both short-term and long-term bone marrow repopulating activity. In addition to myeloid progenitors, this Sca-1+/LIN- population contains a large number of cells that express CD31 and CD34 and have an active Tie-2 promoter, indicating that they are in the endothelial lineage. After 3 to 4 weeks hematopoiesis in treated cultures wanes but if catalase is removed, hematopoiesis resumes. After 7 to 10 days the cultures are indistinguishable from untreated controls. Thus, protected from H2O2, hematopoietic progenitors multiply and become quiescent. This sequence resembles in vivo development in normal marrow. These results make it clear that peroxide-sensitive regulatory mechanisms play an important role in controlling hematopoiesis ex vivo and presumably in vivo as well. They also indicate that manipulation of the peroxide levels can be used to enhance the growth of hematopoietic stem cells in culture.

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Fractionation of mouse bone marrow by adherence separates primitive hematopoietic stem cells from in vitro colony-forming cells and spleen colony-forming cells

Blood ◽

10.1182/blood.v78.10.2577.2577 ◽

1991 ◽

Vol 78 (10) ◽

pp. 2577-2582 ◽

Cited By ~ 22

Author(s):

F Kiefer ◽

EF Wagner ◽

G Keller

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Tissue Culture ◽

Mouse Bone Marrow ◽

Adherent Cells ◽

Tissue Culture Plastic ◽

Adhesive Properties ◽

Hematopoietic Stem

Abstract Fractionation of mouse bone marrow by adherence to tissue culture plastic was used to characterize the adhesive properties of hematopoietic stem (HS) cells capable of long-term reconstitution. The adherent fraction that represents approximately 13% of the total marrow population was virtually devoid of in vitro colony-forming cells and spleen colony-forming cells but did contain approximately 30% of the total HS cells recovered from the procedure. These cells could be detected by both the competitive repopulation assay and by repopulation of W/Wv recipients. In approximately 60% of the recipients from the competitive repopulation experiments, the contribution of the adherent marrow cells was relatively low early (8 to 10 weeks) after transplantation. With time, however, the hematopoietic contribution from these cells increased, reaching a stable level 20 to 30 weeks posttransplantation. In the remaining recipients (40%), the contribution from adherent cells was already significant within 8 to 10 weeks of transplantation and did not change dramatically throughout the course of the experiment. Adherent bone marrow containing significant numbers of HS cells was unable to protect mice from radiation death, indicating that these early cells in the absence of later-stage progenitors are unable to provide this function.

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Hypoxic and normoxic in vitro cultures maintain similar numbers of long-term reconstituting hematopoietic stem cells from mouse bone marrow

Experimental Hematology ◽

10.1016/j.exphem.2012.07.005 ◽

2012 ◽

Vol 40 (11) ◽

pp. 879-881 ◽

Cited By ~ 1

Author(s):

Yanjuan Tang ◽

Camilla Halvarsson ◽

Pernilla Eliasson ◽

Jan-Ingvar Jönsson

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

In Vitro Cultures ◽

Mouse Bone Marrow ◽

Hematopoietic Stem

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SLAM family markers are conserved among hematopoietic stem cells from old and reconstituted mice and markedly increase their purity

Blood ◽

10.1182/blood-2005-05-2140 ◽

2006 ◽

Vol 107 (3) ◽

pp. 924-930 ◽

Cited By ~ 178

Author(s):

Ömer H. Yilmaz ◽

Mark J. Kiel ◽

Sean J. Morrison

Keyword(s):

Stem Cells ◽

Bone Marrow ◽

Hematopoietic Stem Cells ◽

Young Mouse ◽

Receptor Expression ◽

Mouse Bone Marrow ◽

Hematopoietic Stem ◽

Recent Advances ◽

Slam Family

AbstractRecent advances have increased the purity of hematopoietic stem cells (HSCs) isolated from young mouse bone marrow. However, little attention has been paid to the purity of HSCs from other contexts. Although Thy-1lowSca-1+Lineage-c-kit+ cells from young bone marrow are highly enriched for HSCs (1 in 5 cells gives long-term multilineage reconstitution after transplantation into irradiated mice), the same population from old, reconstituted, or cytokine-mobilized mice engrafts much less efficiently (1 in 78 to 1 in 185 cells gives long-term multilineage reconstitution). To test whether we could increase the purity of HSCs isolated from these contexts, we examined the SLAM family markers CD150 and CD48. All detectable HSCs from old, reconstituted, and cyclophosphamide/G-CSF-mobilized mice were CD150+CD48-, just as in normal young bone marrow. Thy-1lowSca-1+Lineage-c-kit+ cells from old, reconstituted, or mobilized mice included mainly CD48+ and/or CD150- cells that lacked reconstituting ability. CD150+CD48-Sca-1+Lineage-c-kit+ cells from old, reconstituted, or mobilized mice were much more highly enriched for HSCs, with 1 in 3 to 1 in 7 cells giving long-term multilineage reconstitution. SLAM family receptor expression is conserved among HSCs from diverse contexts, and HSCs from old, reconstituted, and mobilized mice engraft relatively efficiently after transplantation when contaminating cells are eliminated.

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